1. Field of the Invention
The present invention relates to an apparatus for detecting and measuring the biological activity of microorganisms in a test sample by radiometric techniques and, more specifically, to a test sample container which includes an absorber/scintillation compound positioned relative to the test sample to provide improved measuring efficiency.
2. Prior Art
Radiometric techniques have found wide-spread applications in the biological, medical, food processing and related fields as a method for detecting and measuring biological activity in a test sample. The technique generally involves innoculating a test sample containing or believed to contain bacteria or micro-organisms with a radionuclide or isotope-labeled nutrient media. The micro-organisms metabolize, oxidize, or otherwise process the nutrient media and evolve or respirate a gaseous compound which includes the radioisotope as one of its constituents. Quantitative measurement of this evolved gas and the rate at which it is evolved provides an indication of the presence of and the activity of the microorganisms in the test sample.
The labeled gas metabolically produced by the microorganisms is generally measured by scintillation techniques which involve absorbing the labeled gas in the presence of a scintillation compound or fluor. Light flashes are emitted by the scintillation compound as a function of the quantity of radio-labeled gas that is absorbed. These flashes may then be counted in a conventional laboratory scintillation counter.
A number of often complex laboratory-type devices have been developed for carrying out the above described radiometric techniques. Some of these devices are "one-shot" types in which the experiment must be terminated in order to perform the measurement and are consequently not capable of providing continuous and cumulative text tracking of a sample. Other of these devices require skilled laboratory handling procedures to ensure accurate measurements or cannot be adapted to automatic measurement using scintillation counters which have an automatic sample-vial transport mechanism.
One apparatus, developed by the present inventor, does provide continuous and cumulative tracking and is suitable for use with the automatic sample transports of conventional laboratory scintillation counters. This device is described in an article entitled "Liquid Scintillation Vial for Cumulative and Continuous Radiometric Measurement of In Vitro Metabolism" published by the present inventor in Applied Microbiology, August 1974, pp. 177-180. A similar device is described in U.S. Pat. No. 3,944,471 to Waters. This device includes a first container, e.g., a 30 ml. serum vial, which is adapted to contain the test sample and the nutrient media, and a second container, e.g., a standard scintillation vial, into which the first container is inserted. In one embodiment, a cylindrically formed filter paper, treated with a mixture of an absorber and a scintillation compound, is inserted into the annular space between the outside surface of the serum vial and the inside surface of the scintillation vial. The scintillation vial is then sealed with a gas impermeable closure and the device innoculated with a mixture of the test sample and radio-labeled nutrient-media by means of a hypodermic syringe inserted through the closure. Gas metabolized or fermented as a consequence of the biological activity of the micro-organisms in the test sample fills the available volume between the two containers and is absorbed onto the cylindrical filter paper. The scintillation compound emits light flashes in response to the presence of the radioactive gas absorbed on the filter paper. These flashes, when the vial is mounted in the test well of a conventional laboratory-type scintillation counter, are detected by the counter's photomultiplier tubes and counted to provide a continuous and cumulative quantitative indication of the bacterial activity.
While the above described test sample container provides an acceptable level of measurement efficiency, its structural arrangement is such that a portion of the light flashes emitted by the scintillation compound will not be detected. All conventional liquid scintillation counters use two diametrically opposed photo-multiplier tubes designed to operate in coincidence; that is, a light flash will be registered as a count if, and only if, both tubes simultaneously detect a scintillation flash. The reduction in efficiency can occur, e.g., when a light flash is emitted on one side of the filter paper support with portions of the light energy directed simultaneously towards both photo-multipler tubes. In one case, the light energy passes through the transparent wall of the scintillation vial to be detected by one of the photo-multiplier tubes, and, in the other case, the light energy passes in the opposite direction through the test sample/nutrient media mix that, in many cases, is opaque or only semi-transparent (e.g., blood), causing the light energy emitted toward the second photo-multiplier tube to be absorbed or color quenched in the test sample/nutrient media mix. Another type of quenching, known as chemical quenching, can occur when the light energy impacts long-chain organic molecules in an otherwise transparent test sample/nutrient media mix.
in addition to these count efficiency limitations, the above described design possesses a number of practical drawbacks. The filter paper cylinder and the test sample vial are not secured relative to one another or to the scintillation vial. As a result, it is possible for these two elements to shift position during a test and adversely affect the accuracy of the test and, of course, damage one or the other. The use of cylindrically-formed filter-paper support having a rather large surface area requires that the absorber/scintillation-compound mix be carefully applied over the entire surface of the cylinder to ensure uniform distribution and also requires that a larger than preferably amount of relatively expensive scintillation compound be used with each test sample container.